The physical and enzymatic properties of DNA replication complexes from Escherichia coli and from mammalian cells are being studied to learn details of the normal and semiconservative modes of DNA replication in vivo. The respective roles of DNA polymerases I, II, and III in normal replication and in excision-repair are being examined in toluenized mutant bacterial strains deficient in one or more of these enzymes. The participation of recombination system enzymes in long-patch repair is being studied. Density and radioactive labeling is used to detect repair synthesis and to distinguish it from semiconservative replication by CsCl density gradient sedimentation. Our hypothesis that transcription introduces repairable single strand breaks in bacterial DNA is being tested by selective inhibition of transcription in RNA polymerase deficient mutants or by rifampin, puromycin, or chloramphenicol treatment and subsequent examination of the DNA molecular weight distribution by alkaline sucrose gradient sedimentation. Experiments are being carried out to test the assertion that protein synthesis is a prerequisite for replication of the terminal segment of the E. coli chromosome. Pathways for incorporation and utilization of exogenous thymidine by various strains of E. coli are being studied. This work includes examination of the mechanisms for transport of thymidine through the bacterial membrane (e.g., energetics of thymidine transport). The pathogenic effects of 5-bromouracil in bacteria and in phage lambda are being studied in parallel with an examination of abnormalities in replication and recombination when the analogue is substituted for thymine.